养殖池塘中产毒微囊藻SNP基因型组成与其种群竞争力及产毒特征的关联性分析与应用

Competitive ability and toxin characteristic of toxic Microcystis population are the results of genotypes composition interaction with environmental factor. Furthermore, comparing to environmental factor, genotypes composition, as key endogenous factor, determine the competitive ability and toxin characteristic of toxic Microcystis population. Because of poor discernment of polymorphic sites, existing genotyping techniques could not reveal the correlation among genotype and competitive ability, toxin characteristic of toxic Microcystis population accurately and efficiently. The purpose of present project is to (1) figure out SNP genotypes of toxic Microcystis population based on screening out of all SNP of microcystin biosynthesis gene cluster; (2) figure out the correlation among SNP genotypes composition and competitive ability, toxin characteristic of toxic Microcystis population in aquaculture ponds based on preliminarily grasping the correlation of SNP genotype and competitive ability, toxin characteristic using the experimental ecological method; (3) establish foresight forewarning model of Microcystis blooms harm based on SNP genotypes composition of toxic Microcystis population combining with physical and chemical characteristics of aquaculture waters. The results of present project will provide precise and high-efficient foresight forewarning model of Microcystis blooms harm and reference data for forewarning and emergency prevention and disposal, which has important significance to restoring healthy ecosystem of aquaculture pond, and then achieving the goal of harm reduction of Microcystis bloom to freshwater aquaculture to the most extent.

产毒微囊藻的种群竞争力及产毒特征是其基因型组成与外界环境互作的综合结果，且相比于环境因素的影响作用，基因型组成是决定着微囊藻种群竞争力及产毒特征的关键性内源因素。由于对多态位点识别力差等局限，现有的产毒微囊藻基因分型技术无法准确地揭示产毒藻株基因型与其种群竞争力及产毒特征间关联性。本项目拟在筛查出产毒微囊藻mcy基因簇SNP位点基础上明确产毒微囊藻SNP基因型类别；随后在室内初步掌握SNP基因型与其种群竞争力及产毒特征间关联基础上，明确水华形成过程中养殖池塘内产毒微囊藻SNP基因型组成与其种群竞争力及产毒特征间关联性，结合水体理化指标，建立起基于产毒微囊藻SNP基因型组成的养殖池塘微囊藻水华危害的前瞻性预警模型。研究结果将为养殖池塘产毒微囊藻危害的预警及应急防控工作提供准确高效的预警模型及参考数值，对科学地恢复健康养殖生态系统具有重要意义，进而实现最大程度地降低微囊藻水华对淡水养殖的危害。

产毒微囊藻的种群竞争力及产毒特征是其基因型组成与外界环境互作的综合结果，且相比于环境因素的影响作用，基因型组成是决定着微囊藻种群竞争力及产毒特征的关键性内源因素。本项目在明确养殖池塘MCs时空分布及养殖动物机体MCs积累特征等基础上，研究发现（1）自养殖池塘分离鉴定出若干产毒微囊藻藻株；（2）共培养实验表明产毒微囊藻的种群竞争力非常明显地优于非产毒微囊藻；（3）不同产毒微囊藻藻株产毒量及合成毒素类型差异明显，可划分为6种产毒类型；（3）采用LA-PCR扩增测序获得产毒微囊藻mcy基因簇全长序列，结合其产毒特征可将不同产毒微囊藻藻株初步划分为6种SNP/InDel基因型，具体为MP/MC-RR、MP/MC-LR、HP/MC-RR、LP/MC-RR、HP/MCs、LP/MCs；（4）采用Taqman双标记荧光探针法和Cycling探针法建立起3组两重荧光定量PCR基因分型方法，分型检测特异性和敏感性均较高，同时制备出用于养殖池塘产毒微囊藻基因型定量分析所需的阳性质控标准品；（5）采用项目组建立的荧光定量PCR基因分型方法，原位分析发现对虾养殖池塘产毒微囊藻SNP/InDel基因型随时间变化有明显的演替现象；（6）结合养殖池塘水体理化指标，采用支持向量机回归（SVR）理论和方法建立的预测模型可以很好地模拟养殖池塘中产毒微囊藻SNP/InDel基因型及产毒特征变化规律，模型精度与稳定性均较高。研究结果将为养殖池塘产毒微囊藻危害的预警及应急防控工作提供准确高效的预警模型及参考数值，对科学地恢复健康养殖生态系统具有重要意义，进而实现最大程度地降低微囊藻水华对淡水养殖的危害。